芹菜中转录因子基因AgDREB1的分离与表达研究

Cloning and expression profile analysis of the AgDREB1 transcription factor gene from celery

DREB类转录因子是植物AP2/ERF转录因子家族中一个重要的亚家族,参与植物对逆境的应答,是植物适应逆境的重要调节因子之一。本文从不同品种芹菜中分离DREB类转录因子基因,并研究其表达情况,以进一步研究芹菜中非生物胁迫逆境调控。基于芹菜(Apium graveolens L.)转录组数据,分别从2个芹菜品种‘津南实芹’和‘文图拉’中克隆得到一个编码芹菜DREB类转录因子的基因AgDREB1。采用生物信息学方法对AgDREB1的氨基酸组成、理化性质、进化关系、空间结构等进行了分析。通过实时定量PCR方法对‘津南实芹’和‘文图拉’中AgDREB1基因的表达进行分析。结果表明:来源于‘津南实芹’和‘文图拉’的AgDREB1基因全长分别为495和492 bp,分别编码165和164个氨基酸。芹菜中AgDREB1转录因子等电点约为9.64,包含1个α螺旋和3个β折叠结构,进化关系上属于DREB亚族中的A5组。芹菜AgDREB1转录因子与其他物种的DREB转录因子具有较高的一致性。实时定量PCR分析表明,AgDREB1基因在‘津南实芹’和‘文图拉’根中的表达较高,对低温、高温、干旱和盐胁迫等非生物胁迫有响应。结论:从芹菜中克隆获得一个与逆境相关的DREB类转录因子基因AgDREB1,通过基因表达分析发现,芹菜中AgDREB1基因在根中表达最高,而且该基因与芹菜非生物胁迫相关。

DREB transcription factor is one of the subfamily of AP2/ERF transcription factor family,involving in the responses to biotic and abiotic stresses in higher plant. It is one of the important factors for plant to response to environment stresses. Celery（Apium grave- o/ens L.）is an important biennial vegetable of the Apiaceae family that is widely cultivated around the world. Cetery plants defend themselves from biotic and abiotic stresses by activating various mechanisms involved into transcription regulation. The cDNAs encoding the AgDREB1 gene were cloned from two celery varieties of＇ Jinnanshiqin＇ and＇ Ventura＇ ,respectively. Bioinformaties approaches were utilized to analyze nucleotide and amino acid sequences of the transcription factor of AgDREB1 ,which were cloned from celery（Apium graveolens L. ） cultivars＇ Jinnanshiqin＇ and＇ Ventura＇, respectively. The sequences of DREB transcription factors of higher plants in this study were obtained from the NCBI database. The open reading frames of the AgDREB1 gene were generated by the ORF finder from NCBI. The deduced amino acid sequence, composition, physical and chemical characterization, conserved domain sequence and function domain of the AgDREB1 gene were analyzed. The phylogenetic tree of AgDREB1 transcription factor of celery was created by MEGA 5. The expression profiles of AgDREB1 gene were detected and analyzed by quantitative real-time PCR method under different abiotic stress treatments. The results of sequence analysis showed that lengths of the AgDREB1 gene from the two celery varieties of＇ Jinnan- shiqin＇ and＇ Ventura＇ were 495 and 492 bp,eneeding 165 and 164 amino acids,respectively. The pI of AgDREB1 transcription factor was 9.64. The AgDREB1 transcription factor contained the classic AP2 DNA binding domain, which has the conservative WLG and YRG elements. The AgDREBI of celery was a typical AP2/ERF family transcription factor,which had high similarity with the DREB transcription factors from other plants. Phylogenetic tree created with AP2/ERF family transcription factors of Arabidopsis showed that the AgDREB1 transcription factor of celery was the closest to AtDREB-A5 of Arabidopsis. The AgDREB1 transcription factor of celery was classified into DREB subfamily A5 group. The AgDREB1 transcription factor and AtERF1 （PDB ID： lgcc）had similar three-dimen- sion structure,containing one alpha helix and three beta turn. In the two celery varieties of＇ Jinnanshiqin＇ and＇ Ventura＇ ,quantitative real-time PCR analysis indicated that the AgDREB1 gene was mainly expressed in root ,then in leaf and stem. The AgDREB1 gene was tissue-specific expressed in＇ Jinnanshiqin＇ and＇ Ventura＇. Furthermore, the expression profiles of the AgDREB1 gene under four abiotic stresses（low temperature, high temperature, salt and drought）were detected by quantitative real-time PCR. The gene of AgDREB1 was responsive to cold,hot,drought and high-salt stresses in celery. The gene expression levels of AgDREB1 gene were different in the two celery cuhivars of＇ Jinnanshiqin＇ and ＇ Ventura＇. Conclusions： The AgDREB1 gene, which encodes DREB transcription factor, was cloned from celery. The AgDREB1 gene was tissue-specific expressed in the two celery varieties＇ Jinnanshiqin＇ and＇ Ventura＇ ,and was induced by abiotic stresses in celery.